DESCRIPTION (the Applicant's Abstract): Tuberculosis remains an important clinical problem throughout the world. Protective immunity to tuberculosis depends upon the coordinated response of the cellular immune system. Vaccination strategies that have successfully elicited CD4+ T cells have not been uniformly successful in containing Mtb growth in animal models. Additionally, mice deficient in beta 2 microglobulin and hence MHC class I-dependent T cell responses are exquisitely sensitive to infection with Mtb. These data would argue that CD8+ T cells play an essential and distinct role in the host response to tuberculosis. Identification and characterization of human cytotoxic T cells is important for the understanding of immunity to tuberculosis, and hence may be crucial for the development of efficacious vaccines and improved therapeutic strategies. Work done by the PI and collaborators has previously established the existence of Mtb-specific CD8+ T cells in persons infected with Mtb. Using a sensitive IFN-gamma enzyme linked immunospot (ELISPOT) based limiting dilution analysis (LDA) to evaluate CD8+ T cells grown in response to Mtb-infected dendritic cells (DC), the majority of these cells from one latently-infected individual were found to be non-classically restricted in that they were neither HLA-A, B, or C nor CD 1 restricted. While in the minority, classically (HLA-Ia) restricted cells were also elicited, and the cognate antigen and its minimal peptide epitope defined. Important questions remain unresolved regarding the role of CD8+ T cells in the host response to infection with Mtb in several areas. First, the magnitude of classically restricted non-classically restricted, and CD 1 restricted responses following infection with Mtb is not known. This may be critical in distinguishing a successful (healthy, PPD positive; PPD+) from unsuccessful (tuberculosis) response to infection with Mtb. Second, the molecular basis for antigenic presentation to non-classically restricted CD8+ T cells is not known. Preliminary data are presented indicating that HLA-E may be the restricting molecule for these non-classically restricted cells. Third, prior work has relied upon peptides with predicted binding to HLA motifs to elicit classically restricted CD8+ CTL. One limitation of these peptide-based approaches is that it is difficult to ascertain whether these responses are primed by mycobacterial infection, or represent low-affinity cross-reactivity with another antigen. Similarly, it remains uncertain as to whether the peptides tested reflect dominant epitopes generated during the course of natural infection. Consequently, the question of whether or not antigens known to elicit CD4+ T cell responses also elicit CD8+ T cell responses remains unanswered. Finally, how these Mtb-derived proteins gain access to the HLA-Ia processing pathway remains an important and unresolved question in understanding the human host response to this important pathogen.